TWM655746U - Solid-state culture platform for culturing Antrodia camphorata fruiting body and culture medium device thereof - Google Patents

Abstract

A culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies and a culture medium device thereof mainly include: a culture medium device, the culture medium device having a unit for extracting potato glucose extract, the extractable concentration of which is 2-4% (w/v); a unit for extracting wheat extract, the extractable concentration of which is 2-4% (w/v); a unit for extracting amino acids, the extractable concentration of which is 0.1-0.3% (w/v); and a unit for extracting agar, the extractable concentration of which is 0. 5~0.6% (w/v); an extracted sucrose unit, the extractable concentration of which is 5~7% (w/v), and the solid-state cultured Antrodia cinnamomea fruiting body structure composition produced by combining with the culture medium device includes: Antrodia cinnamomea acid A, Antrodia cinnamomea acid B, Antrodia cinnamomea acid C, Antrodia cinnamomea acid H, Antrodia cinnamomea acid K, dehydrothiochromic acid, dehydroporic acid, 15α-acetyl-dehydrothiochromic acid; wherein the solid-state cultured Antrodia cinnamomea fruiting body structure composition is preserved by the culture platform.

Description

Cultivation platform and culture medium device for solid state culture of Antrodia cinnamomea fruiting bodies

This new type of invention relates to a kind of Antrodia cinnamomea fruiting body, which refers to a solid-state cultured Antrodia cinnamomea fruiting body cultured using a culture medium device, in detail, a culture platform for solid-state cultured Antrodia cinnamomea fruiting body and its culture medium device.

The scientific name of Antrodia cinnamomea is Antrodia cinnamomea, commonly known as Antrodia camphorata, red camphorata, camphor mushroom, camphor mushroom, etc. The fruiting body of Antrodia cinnamomea grows on the inner wall of the decayed heartwood of the trunk of the century-old Antrodia cinnamomum (Cinnamomum kanehirai), or on the dark and moist surface of the dead and fallen Antrodia cinnamomum wood. It is a wood decay fungus. The top surface of Antrodia cinnamomea is brown to dark brown, with unclear wrinkles, glossy, flat and blunt edges, and its ventral surface is orange-red or partially yellow, with many pores.

In addition, Antrodia cinnamomea has a strong sassafras aroma. After drying, it fades to a yellowish-white color and tastes extremely bitter. It is used by the people as a herbal medicine for detoxification, liver protection, and anti-cancer. Antrodia cinnamomea, like other edible and medicinal mushrooms, has many complex ingredients. The known physiologically active ingredients include polysaccharides (such as β-glucan), triterpenoids, superoxide dismutase (SOD), adenosine, protein (including immunoglobulin), vitamins (such as vitamin B, niacin), trace elements (such as calcium, phosphorus and germanium), nucleic acids, lectins, amino acids, sterols, lignin and blood pressure stabilizing substances (such as antodia acid). These physiologically active ingredients are believed to have anti-tumor, immune-enhancing, anti-allergic, platelet aggregation inhibition, anti-viral, anti-bacterial, anti-hypertensive, blood sugar-lowering, cholesterol-lowering and liver protection functions.

Among the many components of Antrodia cinnamomea, triterpenoids are the most studied. Triterpenoids are a general term for natural compounds composed of thirty carbon elements combined into hexagonal or pentagonal shapes. The bitter taste of Antrodia cinnamomea mainly comes from triterpenoids. In 1995, Cherng et al. found that the Antrodia cinnamomea fruiting body extract contained three new triterpenoids with ergostane as the skeleton: antcin A, antcin B and antcin C (Cherng, I.H. and Chiang, H.C. 1995. Three new triterpenoids from Antrodia cinnamomea. J. Nat. Prod. 58: 365-371). In addition, Chiang et al. discovered three other new triterpenoid compounds from the fruiting body extract, namely, antrocin (4,7-dimethoxy-5-methyl-1,3-benzodioxole) and 2,2',5,5'-tetramethoxy-3,4,3',4'-bi-methylenedioxy-6,6'-dimethyl biphenyl) (Chiang, H.C., Wu, D.P., Cherng, I.W. and Ueng, C.H. 1995. A sesquiterpene lactone, phenyl and biphenyl compounds from the fruiting body extract. Antrodia cinnamomea. Phytochemistry. 39: 613-616). In 1996, Cherng et al. discovered four new triterpenoids using the same analytical method: antcin E, antcin F, methyl antcinate G, and methyl antcinate H (Cherng, I.H., Wu, D.P. and Chiang, H.C. 1996. Triteroenoids from Antrodia cinnamomea. Phytochemistry. 41: 263-267). Yang et al. discovered two new compounds with ergostane as the skeleton and three new compounds with lanostane as the skeleton: 15α-acetyl-dehydrosulphurenic acid, dehydroeburicoic acid, and dehydrasulphurenic acid. acid)(Yang,S.W.,Shen,Y.C.and Chen,C.H.1996.Steroids and triterpenoids of Antodia cinnamomea-A fungus parasitic on Cinnamomum micranthum.Phytochemistry.41：1389-1392).

Since wild Antrodia cinnamomea is rare and difficult to obtain, and there is a problem of environmental pollution, in recent years, Antrodia cinnamomea has been cultivated mostly by artificial cultivation methods. Currently, the main cultivation methods for Antrodia cinnamomea are liquid culture method, solid culture method and basswood culture method.

The fruiting bodies of Antrodia cinnamomea have various shapes, such as plate, bell, horseshoe or tower. They are bright red when they are first born, and gradually turn into light reddish brown, light brown or light yellowish brown. Wild white Antrodia cinnamomea is very rare, and it is even rarer to cultivate white Antrodia cinnamomea by artificial cultivation methods. Therefore, white Antrodia cinnamomea is even rarer and more valuable in the market.

In view of the fact that the above-mentioned problems of the prior art cannot be effectively solved and overcome, the present creator, relying on his many years of practical experience and the experience gained from long-term in-depth research and utilization of various Antrodia cinnamomea fruit entities, has proposed this new application to solve the above-mentioned problems.

The main purpose of the present invention is to provide a culture medium device for solid culture of Antrodia cinnamomea fruiting bodies, mainly comprising: a culture medium device, the culture medium device having a unit for extracting potato glucose extract, the extractable concentration of which is 2-4% (w/v); a unit for extracting wheat extract, the extractable concentration of which is 2-4% (w/v); a unit for extracting amino acids, the extractable concentration of which is 0.1-0.3% (w/v); a unit for extracting agar, the extractable concentration of which is 0.5-0.6% (w/v); and a unit for extracting The extractable sucrose unit has an extractable concentration of 5-7% (w/v); wherein the culture medium device is configured with the potato glucose extract unit, the wheat extract unit, the amino acid unit, the agar unit and the sucrose extract unit; the potato glucose extract unit is connected to the wheat extract unit; the wheat extract unit is connected to the amino acid unit; the amino acid unit is connected to the agar unit; the agar unit is connected to the sucrose extract unit.

The secondary purpose of the present invention is to provide a solid-state culture platform for Antrodia cinnamomea fruiting bodies, a white Antrodia cinnamomea fruiting body having a hymenium slice, and the solid-state culture Antrodia cinnamomea fruiting body structure composition produced by the culture medium device includes: cinnamaldehyde A, cinnamaldehyde B, cinnamaldehyde C, cinnamaldehyde H, cinnamaldehyde K, dehydrothiochromic acid, dehydroporic acid, 15α-acetyl-dehydrothiochromic acid; wherein the solid-state culture Antrodia cinnamomea fruiting body structure composition is preserved by the culture platform.

Specifically, the technology used in the present invention is: using a culture platform and a culture medium device for solid-state culture of Antrodia cinnamomea fruiting bodies, mainly including: a culture medium device, the culture medium device has a potato glucose extract unit, the extractable concentration of which is 2~4% (w/v); a wheat extract unit, the extractable concentration of which is 2~4% (w/v); An amino acid extraction unit, whose extractable concentration is 0.1-0.3% (w/v); an agar extraction unit, whose extractable concentration is 0.5-0.6% (w/v); and an sucrose extraction unit, whose extractable concentration is 5-7% (w/v); wherein the culture medium device is configured with the potato glucose extraction unit, the wheat extraction unit, the amino acid extraction unit, the agar extraction unit and the sucrose extraction unit; the potato glucose extraction unit is connected to the wheat extraction unit; the wheat extraction unit is connected to the amino acid extraction unit; the amino acid extraction unit is connected to the agar extraction unit; the agar extraction unit is connected to the sucrose extraction unit, and then a fruiting body is obtained from a white Antrodia cinnamomea fruiting body. The solid-state cultured Antrodia cinnamomea fruiting body structure composition is prepared by cutting into slices and placing them into the culture medium device for culture. The solid-state cultured Antrodia cinnamomea fruiting body structure composition comprises: Antrodia cinnamomea acid A, Antrodia cinnamomea acid B, Antrodia cinnamomea acid C, Antrodia cinnamomea acid H, Antrodia cinnamomea acid K, dehydrothiochromic acid, dehydroporic acid, and 15α-acetyl-dehydrothiochromic acid; wherein the solid-state cultured Antrodia cinnamomea fruiting body structure composition is preserved by the culture platform.

Technical means for solving the problem. To achieve the above-mentioned purpose, the present invention provides a culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies and a culture medium device thereof, which mainly include: a culture medium device, the culture medium device having a potato glucose extract unit, the extractable concentration of which is 2-4% (w/v); a wheat extract unit, the extractable concentration of which is 2-4% (w/v); an amino acid extract unit, the extractable concentration of which is 0.1-0.3% (w/v); an agar extract unit, the extractable concentration of which is 0.5-0.6% (w/v); and an sucrose extract unit, the extractable concentration of which is 5-7% (w/v); wherein the potato glucose extract unit is disposed on the culture medium device. The unit comprises the extracted wheat extract unit, the extracted amino acid unit, the extracted agar unit and the extracted sucrose unit; the extracted potato glucose extract unit is connected to the extracted wheat extract unit; the extracted wheat extract unit is connected to the extracted amino acid unit; the extracted amino acid unit is connected to the extracted agar unit; the extracted agar unit is connected to the extracted sucrose unit, and the solid state cultured Antrodia cinnamomea fruiting body structure composition produced by the culture medium device includes: Antrodia cinnamomea A, Antrodia cinnamomea B, Antrodia cinnamomea C, Antrodia cinnamomea H, Antrodia cinnamomea K, dehydrothiochromic acid, dehydroporic acid, 15α-acetyl-dehydrothiochromic acid; wherein the solid state cultured Antrodia cinnamomea fruiting body structure composition is preserved by the culture platform.

In the embodiment of the present invention, a device for storing anthelmintic acid A is further included, and the anthelmintic acid A is used to inhibit the growth of liver cancer cells.

In the embodiment of the present invention, a device for storing anthelmintic acid B is further included, and the anthelmintic acid B is used to inhibit the growth of liver cancer cells.

In the embodiment of the present invention, a device for storing anthelmintic acid C is further included, and the anthelmintic acid C is used to inhibit the growth of liver cancer cells.

In the embodiment of the present invention, a device for storing anthelmintic acid H is further included, and the anthelmintic acid H is used to inhibit the growth of liver cancer cells.

In the embodiment of the present invention, a device for storing anthelmintic acid K is further included, and the anthelmintic acid K is used to inhibit the growth of liver cancer cells.

In the embodiment of the present invention, a device for storing dehydrothiochromic acid is further included, and the dehydrothiochromic acid can be used to cause mitotic storm and cell apoptosis in human acute myeloid leukemia cells.

In the embodiment of the present invention, a device for storing dehydroporous acid is further included, and the dehydroporous acid is used to reduce the glucose concentration in the blood.

In the embodiment of the present invention, a device for storing 15α-acetyl-dehydrothiochromic acid is further included, and the 15α-acetyl-dehydrothiochromic acid is used to reduce the glucose concentration in the blood.

Through the above description, this new model can obtain the following advantages:

1. The solid cultured Antrodia cinnamomea fruiting bodies can be generated by the culture medium device, which have the same indicative components as the wild red Antrodia cinnamomea and different other components.

2. Effectively improve the production efficiency and preservation of Antrodia cinnamomea fruiting bodies in solid state culture.

3. Solid-state culture of Antrodia cinnamomea fruiting bodies has various medical uses.

100: Culture medium device for solid state culture of Antrodia cinnamomea fruiting bodies

10: Culture medium device

11: Extraction of potato glucose extract unit

12: Extraction of wheat extract unit

13: Extract amino acid units

14: Extraction of agar units

15: Extract sucrose units

200: Solid-state culture platform for Antrodia cinnamomea fruiting bodies

20: Equipment for storing cinnamaldehyde A

21: Equipment for storing cinnamaldehyde B

22: Equipment for storing cinnamaldehyde C

23: Equipment for storing cinnamaldehyde H

24: Equipment for storing cinnamaldehyde K

25: Equipment for storing dehydrothiochromic acid

26: Equipment for storing dehydrogenated porous acid

27: Equipment for storing 15α-acetyl-dehydrothiochromic acid

Figure 1 shows eight known index components and their chemical structures obtained by high performance liquid chromatography determination of the fruiting bodies of Antrodia cinnamomea cultured in solid state in the present invention; Figure 2 shows the results of culturing white Antrodia cinnamomea in three different culture media and measuring the average weight and 95% ethanol extraction rate of each group of white Antrodia cinnamomea; Figure 3 shows the results of stimulating the growth of white Antrodia cinnamomea at two different temperatures and measuring the average weight and 95% ethanol extraction rate of each group of white Antrodia cinnamomea; Figure 4 shows the results of comparing the growth of white Antrodia cinnamomea with and without knife stimulation and measuring the average weight and 95% ethanol extraction rate of each group of white Antrodia cinnamomea. Figure 5 is the result of comparing the growth of white Antrodia cinnamomea with different light intensities, and measuring the average weight and 95% ethanol extraction rate of each group of white Antrodia cinnamomea; Figure 6 is the HPLC chromatogram of the alcohol extract of white Antrodia cinnamomea of the new type; Figure 7 is the HPLC chromatogram of the alcohol extract of Antrodia cinnamomea (red) cultured in basswood on the market; Figure 8 is a block diagram of the culture medium device for solid-state culture of Antrodia cinnamomea fruiting bodies of the new type; Figure 9 is a block diagram of the culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies of the new type.

The new model is described in detail below with the help of the attached drawings and in the form of an embodiment. The drawings used in the text are only for illustration and auxiliary description, and may not be the actual proportion and precise configuration of the new model after implementation. Therefore, the proportion and configuration of the attached drawings should not limit the patent scope of the new model in actual implementation.

Figure 1 shows eight known index components and their chemical structures obtained by high performance liquid chromatography determination of the fruiting bodies of Antrodia cinnamomea cultured in solid state in the present invention. Figure 2 shows the results of culturing white Antrodia cinnamomea in three different culture media and measuring the average weight and 95% ethanol extraction rate of each group of white Antrodia cinnamomea. Figure 3 shows the results of stimulating the growth of white Antrodia cinnamomea at two different temperatures and measuring the average weight and 95% ethanol extraction rate of each group of white Antrodia cinnamomea. Figure 4 shows the results of comparing the growth of white Antrodia cinnamomea with and without knife wound stimulation and measuring the average weight and 95% ethanol extraction rate of each group of white Antrodia cinnamomea. The results of the average weight of Antrodia cinnamomea and 95% ethanol extraction rate, Figure 5 is the result of the new method of stimulating the growth of white Antrodia cinnamomea with different light intensities, and measuring the average weight of each group of white Antrodia cinnamomea and 95% ethanol extraction rate, Figure 6 is the HPLC chromatogram of the alcohol extract of white Antrodia cinnamomea of the new method, Figure 7 is the HPLC chromatogram of the alcohol extract of Antrodia cinnamomea (red) cultured in basswood on the market, Figure 8 is a block diagram of the culture medium device for solid-state culture of Antrodia cinnamomea fruiting bodies, and Figure 9 is a block diagram of the culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies.

As shown in FIG. 1 and FIG. 8 , the culture medium device 100 for solid-state culture of Antrodia cinnamomea fruiting bodies mainly comprises: a culture medium device 10, the culture medium device 10 having a potato glucose extract unit 11, the extractable concentration of which is 2-4% (w/v); a wheat extract unit 12, the extractable concentration of which is 2-4% (w/v); an extracting The culture medium device 10 is provided with the extracting unit 11 of potato glucose extract, the extracting unit 12 of wheat extract, the extracting unit 13 of amino acid, the extracting unit 14 of agar, the extracting unit 15 of sucrose, the extracting unit 16 of sucrose, the extracting unit 17 of sucrose, the extracting unit 18 of sucrose, the extracting unit 19 of sucrose, the extracting unit 20 of sucrose, the extracting unit 21 of sucrose, the extracting unit 22 of sucrose, the extracting unit 23 of sucrose, the extracting unit 24 of sucrose, the extracting unit 25 of sucrose, the extracting unit 26 of sucrose, the extracting unit 27 of sucrose, the extracting unit 28 of sucrose, the extracting unit 29 of sucrose, the extracting unit 20 of sucrose, the extracting unit 21 of sucrose, the extracting unit 27 of sucrose, the extracting unit 28 of sucrose, the extracting unit 29 of sucrose, the extracting unit 29 of sucrose, the extracting unit 20 of sucrose, the extracting unit 21 of sucrose, the extracting unit 22 of sucrose, the extracting unit 23 of sucrose, the extracting unit 24 of sucrose, the extracting unit 25 of sucrose, the extracting unit 26 of sucrose, the extracting unit 27 of sucrose, the extracting unit 28 of sucrose, the extracting unit 29 ... The extracting unit 12, the extracting amino acid unit 13, the extracting agar unit 14 and the extracting sucrose unit 15; the extracting potato glucose extract unit 11 is connected to the extracting wheat extract unit 12; the extracting wheat extract unit 12 is connected to the extracting amino acid unit 13; the extracting amino acid unit 13 is connected to the extracting agar unit 14; the extracting agar unit 14 is connected to The extracted sucrose unit 15 then takes a hymenium slice from a solid-state cultured Antrodia cinnamomea fruiting body and puts it into the culture medium device 10 for culture, so that the solid-state cultured Antrodia cinnamomea fruiting body structure composition produced includes: cinnamaldehyde A, cinnamaldehyde B, cinnamaldehyde C, cinnamaldehyde H, cinnamaldehyde K, dehydrothiochromic acid, dehydroporic acid, 15α-acetyl-dehydrothiochromic acid. Among them, the component indicators detected by the solid-state cultured Antrodia cinnamomea fruiting body meet the national standard CNS16152.

Example 1: Method for obtaining solid-state cultured Antrodia cinnamomea fruiting body slices and purified strains

After disinfecting the blade with 75% alcohol, cut off the white Antrodia cinnamomea mushroom body on the wood and wrap the mushroom body with clean gauze or plastic bag. Before strain separation, wipe the sterile operating table with 75% alcohol and irradiate UV light for 20 minutes, then put the white Antrodia cinnamomea mushroom body into the sterile operating table. When separating, first wipe the surface of the mushroom body with cotton or gauze soaked in 75% alcohol. Use a scalpel to remove the epidermis of the mushroom body, divide the fleshy part inside into 0.5 cm hymenium slices, and inoculate them into a culture dish with 1~3% (w/v) potato dextrose broth (PDB) and 0.5~1.5% (w/v) agar culture medium. Place one piece in each culture dish and cover it with a lid. Place the above culture dish at 20~25℃ and culture it. When the hyphae grow to about 1~3 cm, cut off the outer hyphae and transplant them into a new culture dish. After four to six transplants, confirm that there are no foreign bacteria and you can get the white Antrodia cinnamomea strain.

Example 2: Testing method of weight and 95% ethanol extraction rate.

After the Antrodia cinnamomea fruiting body pieces are harvested, the culture medium is removed and placed in an oven for drying; the setting conditions are 60°C and 200 minutes. After drying, weigh them on a scale; calculate the average weight of the dried pieces. After the dried Antrodia cinnamomea pieces are powdered and mixed evenly, 10g is added to 40 times the volume of 95% ethanol and extracted by ultrasonic vibration for about 1~2 hours. The filtered extract is concentrated and dried by a vacuum concentrator. Weigh the dried extract and divide the weight of the extract by the weight of the original piece to obtain the 95% ethanol extraction rate.

Example 3: Preparation and screening of culture medium

Preparation of white Antrodia cinnamomea culture medium (Formula 1)

The culture medium for solid culture of Antrodia cinnamomea fruiting bodies contains the following ingredients: PDB (Potato Dextrose Broth), with a concentration of 4% (w/v); sucrose, with a concentration of 5% (w/v); amino acids, with a concentration of 0.3% (w/v); agar, with a concentration of 0.5% (w/v). The nitrogen source is not limited to amino acids. After the components are mixed, place them in one or more sterilizable containers and add reverse osmosis water to mix and dissolve. Then place them in a sterilizer; set the conditions to 121℃, 15~30 minutes. Pour the sterilized culture medium solution into the culture dish, about 12~27ml per culture dish. After cooling, a solid culture medium can be obtained. The separated white Antrodia cinnamomea strains are implanted in the culture medium and cultured in an environment of 22-30℃ for 180 days.

Preparation of white Antrodia cinnamomea culture medium (Formula 2)

The culture medium for solid state culture of Antrodia cinnamomea fruiting bodies contains the following ingredients: PDB (Potato Dextrose Broth), with a concentration of 2% (w/v); Malt extract, with a concentration of 2% (w/v); sucrose, with a concentration of 7% (w/v); amino acids, with a concentration of 0.2% (w/v); agar, with a concentration of 0.6% (w/v). After the components are mixed, they are placed in one or more sterilizable containers and reverse osmosis water is added to mix and dissolve. Then they are placed in a sterilization autoclave; the setting conditions are 121°C and 15 to 30 minutes. Pour the sterilized culture medium solution into culture dishes, about 15 to 25 ml per culture dish. After cooling, a solid culture medium can be obtained. The separated white Antrodia cinnamomea strains are implanted in the culture medium and cultured in an environment of 22-30℃ for 180 days.

Preparation of white Antrodia cinnamomea culture medium (Formula 3)

The culture medium for solid culture of Antrodia cinnamomea fruiting bodies contains the following ingredients: Malt extract, with a concentration of 4% (w/v); sucrose, with a concentration of 6% (w/v); amino acids, with a concentration of 0.1% (w/v); agar, with a concentration of 0.5% (w/v). After the components are mixed, they are placed in one or more sterilizable containers and reverse osmosis water is added to mix and dissolve. Then they are placed in a sterilization autoclave; the setting conditions are 121°C and 15 to 30 minutes. Pour the sterilized culture medium solution into culture dishes, about 15 to 25 ml per culture dish. After cooling, the solid culture medium is obtained. The isolated white Antrodia cinnamomea strains were implanted into the culture medium and cultured in an environment of 22-30°C for 180 days.

The test method described in Example 2 was used to measure the weight of the fruiting bodies and the 95% ethanol extraction rate obtained after 180 days of culture for the three culture medium formulas mentioned above; the results are shown in Figure 2.

As shown in Figure 2, among the solid-state cultured Antrodia cinnamomea fruiting bodies grown by three different formulas, the first PDB formula (Formula 1) has an average dry weight of 0.9g and a 95% ethanol extraction rate of 28%. The second formula PDB plus wheat extract (Formula 2); the average dry weight is 1.18g, and the 95% ethanol extraction rate is 30%. The third formula wheat extract (Formula 3); the average dry weight is 0.85g, and the 95% ethanol extraction rate is 25%. It shows that the ingredients of Formula 2 are better than Formula 1 and Formula 3.

Example 4: Low temperature stimulation cultivation of white Antrodia cinnamomea growth environment regulation

The best quality medium (Formula 2) was selected from the three culture media described in Example 3 for low temperature stimulation culture. Three groups of culture dishes with white Antrodia cinnamomea in the culture medium (Formula 2) were placed in an environment of 22-30°C for 90 days. The first group was the control group; the culture was continued for a total of 180 days before harvesting and drying. The other two groups of culture dishes were placed in an environment of 4°C and 15°C for 3-5 days respectively, and then the culture dishes were returned to an environment of 22-30°C until a total of 180 days before harvesting and drying. The fruiting bodies obtained under three different culture conditions were measured for weight and 95% ethanol extraction rate using the detection method described in Example 2. The results are shown in Figure 3.

The results in Figure 3 are different low temperature stimulations. The first group is 15℃; the second group is 4℃. The results show that the average dry weight of the first group is 1.18g, and the 95% ethanol extraction rate is 33%; the average dry weight of the second group is 1.18g, and the 95% ethanol extraction rate is 36%. It shows that the low temperature stimulation culture weight remains unchanged, but the ethanol extraction rate increases by 1% and 2% respectively; and the effect of 4℃ is higher than the result obtained at 15℃.

Example 5: White Antrodia camphorata growth environment regulation - knife wound stimulation cultivation

The medium with better quality (formula 2) among the above three mediums was used for stimulation culture. After the culture dishes with white Antrodia cinnamomea inoculated in the culture medium were cultured in an environment of 22-30℃ for 90 days, one group (stimulation group) of culture dishes with white Antrodia cinnamomea grown was taken out, and 0.5-1 cm scars were made on the white Antrodia cinnamomea with a blade or needle to simulate insect bites to stimulate the synthesis of secondary metabolites. After that, they were returned to an environment of 22-30℃ and cultured for 180 days. The other group (non-stimulation group) was cultured in an environment of 22-30℃ for 180 days. The weight and 95% ethanol extraction rate of the solid cultured Antrodia cinnamomea fruiting bodies obtained from the two groups were measured by the detection method described in Example 2, and the results are shown in Figure 4.

Figure 4 shows the results of the knife-wound stimulation experiment. The first group was the group without knife-wound stimulation, and the average dry weight and 95% ethanol extraction rate were 1.2g and 30% respectively. The second group was the group with knife-wound stimulation; the average dry weight and 95% ethanol extraction rate of the mushroom body stimulated by the wound were 1.53g and 35% respectively, which increased by 27% and 16% respectively compared with the first group without knife-wound experiments. It shows that the stimulation of knife-wounds helps the synthesis of secondary metabolites and the increase of weight.

Example 6: Cultivation of white Antrodia cinnamomea stimulated by different light intensities

The best quality medium (Group 2) was selected from the three culture media described in Example 4. The culture dish with the white Antrodia cinnamomea strain implanted in the culture medium was placed in an environment of 22-30°C for 75 days before light stimulation. This experiment was divided into five groups, namely the control group (no light culture), the first group (red light) with a light intensity of 20 μmol/S.m2, the second group (red light) with a light intensity of 12 μmol/S.m2, the third group (blue light) with a light intensity of 16 μmol/S.m2, and the fourth group (blue light) with a light intensity of 8 μmol/S.m2. Except for the control group, each group was stopped from being irradiated after 15 days of irradiation, and was placed in an environment of 22-30°C for 180 days. The weight and 95% ethanol extraction rate of the five groups of solid-state cultured Antrodia cinnamomea fruiting bodies were measured using the detection method described in Example 2. The results are shown in Figure 5.

Figure 5 is a light stimulation experiment. The average weight of the control group was 1.2g; the 95% ethanol extraction rate was 30%. The average weights of the first to fourth groups of light stimulation were 1.18g, 1.16g, 1.17g, and 1.19g, respectively, which were not much different from the control group. In the results of light stimulation at 95% ethanol extraction rate, the average 95% ethanol extraction rates of the first to fourth groups were 36%, 32.5%, 34.8%, and 31.5%, respectively. Compared with the 95% ethanol extraction rate of the control group, they increased by 20%, 8.3%, 16%, and 5%, respectively. The results showed that the average weight of the groups with light stimulation was not much different, while the 95% ethanol extraction rate increased. Among them, the ethanol extraction rate obtained with a red light intensity of 20μmol/S.m2 was the highest.

Example 7: Preparation method of Antrodia cinnamomea alcohol extract

Take 3 grams of commercially available basswood cultured Antrodia cinnamomea fruiting bodies (red) and fresh white Antrodia cinnamomea cultured in Example 4, freeze-dry them in a freeze dryer (temperature below -45°C, vacuum below 300mTorr). The dried products are then crushed in a grinder and poured into serum bottles, and 10 times the volume of 95% alcohol and a stirrer are added, and the mixture is stirred and extracted for 3-5 days by cold soaking. Vacuum filter (No.1 filter paper), concentrate, dry and weigh, which is the Antrodia cinnamomea alcohol extract, and store it in a moisture-proof box.

Example 8: Fingerprint analysis of white Antrodia cinnamomea extract

1 The Antrodia cinnamomea extract prepared in Example 6 was re-dissolved in methanol to a concentration of 5 mg/mL.

2 Centrifuge, 10000rpm, 10mins, take the supernatant for HPLC analysis.

3 High performance liquid chromatography analyzer: Pump: Spectra SYSTEM P1000, Auto injector: Spectra SYSTEM AS1000, Detector: FINNIGAN SURVEYOR PDA Plus Detector

Liquid chromatography analysis conditions:

Chromatographic column: Agilent, Eclipse XDB-C18, 4.6mm*150mm, 5μm

Detector: PDA (UV 254nm)

Sample injection volume: 5μL

Chromatographic flow rate: 0.8mL/min

Withdrawal conditions (as shown in Table 1):

The analysis results of the white Antrodia cinnamomea extract are shown in Figure 6, and the commercially available Antrodia cinnamomea fruiting body (red) alcohol extract cultivated on basswood is shown in Figure 7. Comparison of the HPLC chromatograms of Figures 6 and 7 and the reference (Taiwan-endemic Antrodia cinnamomea (mushroom) (Antrodia cinnamomea) fruiting body's latest standard and correct scientific name, 2013.08) and the Taiwan Edible and Medicinal Mushroom Biotechnology Association published HPLC chromatograms showing the characteristic chromatographic peaks of Antrodia cinnamomea fruiting bodies cultured in basswood. It can be clearly seen that the white Antrodia cinnamomea extract has the characteristic chromatographic peaks of Antrodia cinnamomea fruiting bodies cultured in basswood (as shown in Table 2). In addition, the white Antrodia cinnamomea extract also has some chromatographic peaks different from the Antrodia cinnamomea fruiting bodies cultured in basswood (as shown in Figure 6, the retention times are 4.2, 6.5, 6.8, 12.7 and 20.5 min, etc.), indicating that the white Antrodia cinnamomea obtained by the present invention has more diverse components.

As shown in Figures 1, 8 and 9, the present invention provides a culture platform 200 for solid-state culture of Antrodia cinnamomea fruiting bodies, a culture medium device 100 for solid-state culture of Antrodia cinnamomea fruiting bodies, and a white Antrodia cinnamomea fruiting body having a hymenium slice, and the solid-state cultured Antrodia cinnamomea fruiting body structure composition produced by the culture medium device 10 includes: cinnamaldehyde A, cinnamaldehyde B, cinnamaldehyde C, cinnamaldehyde H, cinnamaldehyde K, dehydrothiochromatic acid, dehydroporic acid, and 15α-acetyl-dehydrothiochromatic acid; wherein the solid-state cultured Antrodia cinnamomea fruiting body structure composition is preserved by the culture platform 200. The culture platform 200 for solid state culture of Antrodia cinnamomea fruiting bodies further includes a device 20 for storing antinotriac acid A, and the antinotriac acid A (Antcin A) is used to inhibit the growth of liver cancer cells; further includes a device 21 for storing antinotriac acid B, and the antinotriac acid B (Antcin B) is used to inhibit the growth of liver cancer cells; further includes a device 22 for storing antinotriac acid C, and the antinotriac acid C (Antcin C) is used to inhibit the growth of liver cancer cells; further includes a device 23 for storing antinotriac acid H, and the antinotriac acid H (Antcin H) is used to inhibit the growth of liver cancer cells; further includes a device 24 for storing antinotriac acid K, and the antinotriac acid K (Antcin K), which is used to inhibit the growth of liver cancer cells; it also includes a device 25 for storing dehydrosulphurenic acid, and the dehydrosulphurenic acid (DSA) can be used to cause human acute myeloid leukemia cells to produce mitotic storms and cell apoptosis; it also includes a device 26 for storing dehydroeburicoic acid, and the dehydroeburicoic acid (DEA) is used to reduce the glucose concentration in the blood; it also includes a device 27 for storing 15α-acetyl dehydrosulphurenic acid, and the 15α-acetyl dehydrosulphurenic acid (15α-Acetyl DSA) is used to reduce the glucose concentration in the blood.

Through the above description, the new type can generate solid cultured Antrodia cinnamomea fruiting bodies through the culture medium device, which has the same indicative components as wild red Antrodia cinnamomea and different other components, which meets the needs of progress, practicality and users, and its benefits are evident.

The feature of this new method is that the solid-state cultured Antrodia cinnamomea fruiting bodies have various medical uses.

In summary, this new type of structure has never been seen in any books or public use, and it truly meets the requirements for applying for a new type of patent. I sincerely request the Bureau to examine it and approve the patent as soon as possible. I would be very grateful.

It should be noted that the above is the technical principle used in the specific implementation of this new model. If the changes made according to the concept of this new model still do not exceed the spirit covered by the instructions and drawings, they should be stated within the scope of this new model.

100: Culture medium device for solid state culture of Antrodia cinnamomea fruiting bodies

10: Culture medium device

200: Solid-state culture platform for Antrodia cinnamomea fruiting bodies

20: Equipment for storing antrodia cinnamomea A

21: Equipment for storing cinnamaldehyde B

22: Equipment for storing cinnamaldehyde C

23: Equipment for storing cinnamaldehyde H

24: Equipment for storing cinnamaldehyde K

25: Equipment for storing dehydrothiochromic acid

26: Equipment for storing dehydrogenated porous acid

27: Equipment for storing 15α-acetyl-dehydrothiochromic acid

Claims (10)

A culture medium device for solid-state culture of Antrodia cinnamomea fruiting bodies is characterized in that it mainly comprises: a culture medium device, the culture medium device having a unit for extracting potato glucose extract, whose extractable concentration is 2-4% (w/v); a unit for extracting wheat extract, whose extractable concentration is 2-4% (w/v); a unit for extracting amino acids, whose extractable concentration is 0.1-0.3% (w/v); a unit for extracting agar, whose extractable concentration is 0.5-0.6% (w/v); a unit for extracting sugarcane extract, whose extractable concentration is 0.1-0.3% (w/v); a unit for extracting sucrose, whose extractable concentration is 0.5-0.6% (w/v); a unit for extracting sucrose ... Sugar unit, the extractable concentration of which is 5-7% (w/v); wherein the culture medium device is configured with the potato glucose extract unit, the wheat extract unit, the amino acid unit, the agar unit and the sucrose unit; the potato glucose extract unit is connected to the wheat extract unit; the wheat extract unit is connected to the amino acid unit; the amino acid unit is connected to the agar unit; the agar unit is connected to the sucrose unit. A culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies, characterized by a culture medium device, the culture medium device having a potato glucose extract unit, the extractable concentration of which is 2-4% (w/v); a wheat extract unit, the extractable concentration of which is 2-4% (w/v); an amino acid extract unit, the extractable concentration of which is 0.1-0.3% (w/v); an agar extract unit, the extractable concentration of which is 0.5-0.6% (w/v); and a sucrose extract unit, the extractable concentration of which is 5-7% (w/v); wherein the potato glucose extract unit, the wheat extract unit, and the sucrose extract unit are arranged on the culture medium device. , the extracted amino acid unit, the extracted agar unit and the extracted sucrose unit; the extracted potato glucose extract unit is connected to the extracted wheat extract unit; the extracted wheat extract unit is connected to the extracted amino acid unit; the extracted amino acid unit is connected to the extracted agar unit; the extracted agar unit is connected to the extracted sucrose unit, and a white Antrodia cinnamomea fruiting body has a hymenium slice, and the solid-state cultured Antrodia cinnamomea fruiting body structure composition produced by combining the culture medium device includes: Antrodia cinnamomea A, Antrodia cinnamomea B, Antrodia cinnamomea C, Antrodia cinnamomea H, Antrodia cinnamomea K, dehydrothiochromic acid, dehydroporic acid, 15α-acetyl-dehydrothiochromic acid. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for preserving antrodia cinnamomea acid A, and the antrodia cinnamomea acid A is used to inhibit the growth of liver cancer cells. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for preserving antrodia acid B, and the antrodia acid B is used to inhibit the growth of liver cancer cells. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for preserving anthelmintic acid C, and the anthelmintic acid C is used to inhibit the growth of liver cancer cells. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for preserving antrodia cinnamomea H, and the antrodia cinnamomea H is used to inhibit the growth of liver cancer cells. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for preserving cinnamomea acid K, and the cinnamomea acid K is used to inhibit the growth of liver cancer cells. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for storing dehydrothiochromic acid, and the dehydrothiochromic acid can be used to cause mitotic storm and cell apoptosis in human acute myeloid leukemia cells. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for storing dehydroporous acid, and the dehydroporous acid is used to reduce the glucose concentration in the blood. The culture platform for solid-state culture of Antrodia cinnamomea fruiting bodies as described in claim 2 further includes a device for preserving 15α-acetyl-dehydrothiochromoporic acid, and the 15α-acetyl-dehydrothiochromoporic acid is used to reduce the glucose concentration in the blood.